title: Characterization of molecular and cellular functions of the cyclin-dependent kinase CDK9 using a novel specific inhibitor.
abstract: OBJECTIVE
  The cyclin-dependent kinase CDK9 is an important therapeutic target but currently available inhibitors exhibit low specificity and/or narrow therapeutic windows. Here we have used a new highly specific CDK9 inhibitor, LDC000067 to interrogate gene control mechanisms mediated by CDK9.
  
  
  METHODS
  The selectivity of LDC000067 was established in functional kinase assays. Functions of CDK9 in gene expression were assessed with in vitro transcription experiments, single gene analyses and genome-wide expression profiling. Cultures of mouse embryonic stem cells, HeLa cells, several cancer cell lines, along with cells from patients with acute myelogenous leukaemia were also used to investigate cellular responses to LDC000067.
  
  
  RESULTS
  The selectivity of LDC000067 for CDK9 over other CDKs exceeded that of the known inhibitors flavopiridol and DRB. LDC000067 inhibited in vitro transcription in an ATP-competitive and dose-dependent manner. Gene expression profiling of cells treated with LDC000067 demonstrated a selective reduction of short-lived mRNAs, including important regulators of proliferation and apoptosis. Analysis of de novo RNA synthesis suggested a wide ranging positive role of CDK9. At the molecular and cellular level, LDC000067 reproduced effects characteristic of CDK9 inhibition such as enhanced pausing of RNA polymerase II on genes and, most importantly, induction of apoptosis in cancer cells.
  
  
  CONCLUSIONS
  Our study provides a framework for the mechanistic understanding of cellular responses to CDK9 inhibition. LDC000067 represents a promising lead for the development of clinically useful, highly specific CDK9 inhibitors.
authors.affiliation: Institute of Molecular Tumor Biology (IMTB), Faculty of Medicine, Westfalian Wilhelms University Muenster (WWU), Muenster, Germany.
authors.forename: T K
authors.initials: TK
authors.lastname: Albert
authors.forename: C
authors.initials: C
authors.lastname: Rigault
authors.forename: J
authors.initials: J
authors.lastname: Eickhoff
authors.forename: K
authors.initials: K
authors.lastname: Baumgart
authors.forename: C
authors.initials: C
authors.lastname: Antrecht
authors.forename: B
authors.initials: B
authors.lastname: Klebl
authors.forename: G
authors.initials: G
authors.lastname: Mittler
authors.forename: M
authors.initials: M
authors.lastname: Meisterernst
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D049109:Cell Proliferation / Q000187:drug effects; D042863:Cyclin-Dependent Kinase 9 / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D045325:HCT116 Cells; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D061986:MCF-7 Cells; D051379:Mice; D035683:MicroRNAs / Q000378:metabolism; D058990:Molecular Targeted Therapy; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D011743:Pyrimidines / Q000494:pharmacology*; D012319:RNA Polymerase II / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013449:Sulfonamides / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection
chemical_list: C000590771:3-((6-(2-methoxyphenyl)pyrimidin-4-yl)amino)phenyl)methane sulfonamide; D035683:MicroRNAs; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D012333:RNA, Messenger; D013449:Sulfonamides; D000255:Adenosine Triphosphate; C467969:CDK9 protein, human; D042863:Cyclin-Dependent Kinase 9; D012319:RNA Polymerase II
keywords: CDK9; RNA polymerase II; cyclin-dependent kinase; gene expression; transcription